Circadian Rhythms and Sleep in Drosophila melanogaster

Abstract

The advantages of the model organism Drosophila melanogaster, including low genetic redundancy, functional simplicity, and the ability to conduct large-scale genetic screens, have been essential for understanding the molecular nature of circadian (∼24 hr) rhythms, and continue to be valuable in discovering novel regulators of circadian rhythms and sleep. In this review, we discuss the current understanding of these interrelated biological processes in Drosophila and the wider implications of this research. Clock genes period and timeless were first discovered in large-scale Drosophila genetic screens developed in the 1970s. Feedback of period and timeless on their own transcription forms the core of the molecular clock, and accurately timed expression, localization, post-transcriptional modification, and function of these genes is thought to be critical for maintaining the circadian cycle. Regulators, including several phosphatases and kinases, act on different steps of this feedback loop to ensure strong and accurately timed rhythms. Approximately 150 neurons in the fly brain that contain the core components of the molecular clock act together to translate this intracellular cycling into rhythmic behavior. We discuss how different groups of clock neurons serve different functions in allowing clocks to entrain to environmental cues, driving behavioral outputs at different times of day, and allowing flexible behavioral responses in different environmental conditions. The neuropeptide PDF provides an important signal thought to synchronize clock neurons, although the details of how PDF accomplishes this function are still being explored. Secreted signals from clock neurons also influence rhythms in other tissues. SLEEP is, in part, regulated by the circadian clock, which ensures appropriate timing of sleep, but the amount and quality of sleep are also determined by other mechanisms that ensure a homeostatic balance between sleep and wake. Flies have been useful for identifying a large set of genes, molecules, and neuroanatomic loci important for regulating sleep amount. Conserved aspects of sleep regulation in flies and mammals include wake-promoting roles for catecholamine neurotransmitters and involvement of hypothalamus-like regions, although other neuroanatomic regions implicated in sleep in flies have less clear parallels. Sleep is also subject to regulation by factors such as food availability, stress, and social environment. We are beginning to understand how the identified molecules and neurons interact with each other, and with the environment, to regulate sleep. Drosophila researchers can also take advantage of increasing mechanistic understanding of other behaviors, such as learning and memory, courtship, and aggression, to understand how sleep loss impacts these behaviors. Flies thus remain a valuable tool for both discovery of novel molecules and deep mechanistic understanding of sleep and circadian rhythms.

Keywords: FlyBook: Drosophila; circadian rhythms; molecular neuroscience; neuroscience; sleep.

Figure 1

(A) Activity for a group of wild-type (WT) male flies in a 12:12 hr light:dark (LD) cycle at 25°. Flies anticipate lights-off, and under these conditions also lights-on, with increased activity in advance of these transitions. (B) Double-plotted activity in constant darkness for two individual WT and per⁰¹ male flies after entrainment in standard LD conditions. Data are double-plotted for ease of interpretation, with each day of data plotted in a new row concatenated with the data from the subsequent day, such that Row 1 displays data for Day 1 and Day 2, Row 2 displays data for Day 2 and Day 3, etc. Activity is concentrated in the subjective day in a pattern that recurs with a period of slightly <24 hr in WT flies.

Figure 2

The molecular feedback loop is formed by the negative feedback of Period (PER) and Timeless (TIM) on their own transcription. Delays exist between transcription of per and tim mRNA and the localization of these proteins in the nucleus, where they can interact with transcriptional activators Clock (CLK) and Cycle (CYC). These delays are thought to be important for allowing the molecular clock to cycle with a period of ∼24 hr. Critical regulators have been identified at several steps of the cycle that are necessary for accurate timing and strength of molecular rhythms. Degradation of PER and TIM allows the cycle to start anew. Not pictured is the second feedback loop formed by PDP1 and Vrille, which produces cycling of Clk mRNA. This secondary feedback loop is thought to reinforce molecular oscillations, although cycling of the CLK protein is not necessary for rhythms. CKII, Casein Kinase II; SGG, shaggy; PP2A, Protein Phosphatase 2A; PP1, Protein Phosphatase 1; DBT, doubletime.

Figure 3

Clock cells, which express the core components of the molecular clock, are depicted on the right. These cells are interconnected and heterogeneous between and within clusters, allowing cells to serve different functions, and respond to different environmental conditions. On the left, two groups of output neurons that do not express the molecular components of the clock, but have cycling neuronal activity and are important for behavioral activity rhythms, suggesting clock input. DN, dorsal neurons; LN, lateral neurons (lLNv, large ventral lateral neurons; sLNv, small ventral lateral neurons; LNd, dorsal lateral neurons); LPN, lateral posterior neurons; LHLK, lateral horn leucokinin neuron; DH44, Diuretic Hormone 44.

Figure 4

(A) Sleep behavior for a group of wild-type (WT) female flies in a 12:12 hr light:dark cycle. Flies have short bouts of siesta sleep in the middle of the day (more pronounced in males) and a relatively consolidated period of sleep at night. (B) Sleep behavior for WT and per⁰¹ male flies in constant darkness (DD). per⁰¹ flies, which do not display circadian rhythms of activity, spend approximately the same amount of time in sleep, but have sleep that is fragmented across the day. Data appear slightly noisier as fewer flies are represented compared to (A).

Figure 5

(A) Schematic of sleep-promoting (red), and sleep-inhibiting (blue), neurons in the fly brain. Sleep-regulating neurons are identified by neurotransmitter, neuropeptide, or molecular marker expression, and/or neuroanatomic location. Dopaminergic neurons: PAM, protocerebral anterior lateral; PPL1, protocerebral posterior lateral; and PPM3, protocerebral posterior medial. Mushroom body (MB) neurons: KC, Kenyon cells; MBON, mushroom body output neurons. Central complex: dFSB, dorsal fan-shaped body; EB, ellipsoid body. Pars intercerebralis (PI): SIFaR, SIFamide Receptor; Rho, Rhomboid; and dILP, Drosophila insulin-like peptide. Octopaminergic neurons: ASM, anterior superior medial. Pars lateralis (PL): CycA, CyclinA. Clock cells: DN, dorsal neurons; lLNvs, large ventral lateral neurons. (B) Location of sleep-regulating neurons in the fly brain.